This invention relates generally to novel synthetic polynucleotides which have been chemically modified by the introduction of reactive mercapto groups and also fluorine atoms at the 5-positions of some of the cytosine and uracil bases in their polymeric chains. The partially unsubstituted polynucleotides are useful in blocking the function of nucleic acid polymerases which are essential for the replication of neoplastic mammalian cells. In addition to being potent inhibitors of nucleic acid polymerases, complexes containing the partially substituted polynucleotides also actively induce synthesis of interferon.
Nucleic acid polymerases have been shown to be target enzymes of several anti-cancer drugs. For example, Bardos et al, Vol. 13, page 359, Proceedings of American Association of Cancer Research, first proposed the use of modified polynucleotides containing mercapto groups at the 5-position of some of the pyrimidine bases of the polymeric molecules as inhibitors of nucleic acid polymerases. In general, the early study showed that partially thiolated DNA and RNA isolates, 5-mercaptopolycytidylic acid (MPC) and 5-mercaptopoly-uridylic acid (MPU), were useful as inhibitors of DNA-dependent RNA polymerases, RNA-directed DNA polymerases (reverse transcriptases) and DNA polymerase alpha from regenerating (normal) rat liver.
In addition to the inhibitory antitemplate activity of partially thiolated polynucleotides in a DNA polymerase system, i.e., selective and strong binding to a specific target enzyme in a manner to prevent binding of the natural template or primer required for gene replication or transcription and cell growth, double-stranded polynucleotide complex, poly(inosinic acid)-5-mercaptopolycytidylic acid (PolyI.MPC) has shown to be capable of inducing human alpha, beta and gamma interferons. Both the inhibitory (antitemplate) and interferon inducing properties of such polynucleotide may contribute to its overall favorable antitumor activity.
In contrast to the antitemplate mode of action of partially thiolated polynucleotides, halogenated polynucleotides, such as fluorinated polycytidylic acid (FPC), were found to lack DNA polymerase inhibitory activity. Instead, they appear to exert their cytotoxic effects via some other mechanism. Thus, even though FPC lacks the desired selectivity associated with antitemplate inhibitory polynucleotides it nevertheless has been shown to be actually more cytotoxic than MPC in some in vitro cell culture systems, presumably due to the cytotoxic effect of the monomeric 5-fluorocytidine liberated as a result of the hydrolysis of FPC.
In view of the observation that base compositions of modified heteropolynucleotides appear to have profound effects on the mode of action of antitumor compounds, it would be highly desirable to have compounds which provide a broader spectrum of antitumor activities, such as acting as potent cytoxic agents while also inhibiting replication of tumor cells through the antitemplate mode of action, including those which act on the immunologic system. Accordingly, the present invention provides for novel partially substituted polynucleotides, methods for actively inhibiting the replication of neoplastic cells in the in vivo biological system, including methods for inducing the synthesis of interferon.